The prevalence of diabetes and heart diseases is increasing in the world. Nutraceuticals of natural origin are gaining importance as an alternative to modern drugs for the management of metabolic syndrome. In the present study, punicic acid (PA), a major bioactive found in pomegranate seed, was subjected for biological characterization with respect to peroxisome proliferator-activated receptor gamma (PPARγ) agonist property in an in vitro system (3T3-L1 adipocytes). We evaluated the adipogenic potential of various concentrations (5, 10 and 30 μM) of PA by studying triglyceride accumulation and glycerol-3-phosphate dehydrogenase (GPDH) activity in adipocytes, which were found to be increased moderately compared with the positive control, i.e. rosiglitazone (RG). Glucose uptake activity (↑225.93% ± 2.55% for 30 μM of PA), and the prevention of reactive oxygen species (ROS) generation (↓57 ± 1.83% for 30 μM of PA) in adipocytes with PA were also evaluated. We also found that PA increased adiponectin secretion and upregulated GLUT4 expression and translocation in adipocytes. Molecular modelling studies revealed a high binding affinity of PA to the PPARγ ligand binding domain. An in vitro ligand binding assay based on time-resolved fluorescence resonance energy transfer (TR-FRET) also proved PA as a PPARγ agonist. Finally, we conclude that PA is a potential nutraceutical and should be encouraged for use both as a prophylactic and therapeutic agent.